The primary objective of this project is to determine the mechanism by which chemical carcinogens such as 3,4-benzpyrene and 2- acetylaminofluorene are activated and destroyed by the microsomal mixed- function oxidase system present in human tissues such as the liver, lung, and skin. Cytochrome P-450 plays an essential role in the oxygenation reactions which appear to be the first step in the metabolism of most carcinogens. Chloroperoxidase exhibits striking structural and functional similarities to cytochrome P-540, therefore, it will be used as a model system for the study of the mechanisms of oxygen activation and substrate oxygenation by the liver microsomal system since it is a pure protein which is well characterized and readily available. The initial phases of experimentation will be concerned with studies on the various types of oxygenation reactions catalyzed by this enzyme. These reactions will be investigated with respect to types of oxygen utilized, substrate specificity, and identity of products. The binding of carcinogenic substrates and other ligands to chloroperoxidase will be investigated by optical, ESR, and NMR spectroscopy in order to learn more about the substrate binding sites, the interaction of substrates with the heme-iron and the porphyrin ring, and the interaction of various substrates with enzyme-bound oxygen. The interaction between the protein and various substrates will also be investigated by measuring the effect of ligand binding on the redox potentials of the heme iron. Amino acid modification studies using a variety of specific reagents will be carried out in an attempt to identify the axial ligand which is responsible for the unusual spectral and catalytic properties of P-450 type cytochromes and the role of the heme ligands in catalysis.